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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

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Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
28 May 2018 - 27 Jun 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Cross-reference
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
2004
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
Appearance: Clear colourless to pale yellow liquid
Test item storage: In refrigerator (2-8°C)

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: Control and all test concentrations
- Sampling method: 2.0 mL taken from the approximate centre of the test vessels, at t=0 h and t=48 h.
- At the end of the exposure period, the replicates were pooled at each concentration before sampling.
- Sample storage conditions before analysis: Samples were stored in a freezer (≤-15°C) until analysis at the analytical laboratory of the Test Facility.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Direct addition to the test medium. Test solutions were prepared at the highest concentration of 100 mg/L applying an overnight period of magnetic stirring to accelerate dissolution of the test item in medium. Lower test concentrations were prepared by subsequent dilutions of the highest concentration in test medium. All test solutions were clear and colorless at the end of the preparation procedure.
- Controls: Test medium without test item or other additives.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: Straus 1820
- Source: In-house laboratory culture with a known history
- Age of parental stock: >2 weeks old
- Age at test initiation: <24 hours
- Validity of batch: Daphnids originated from a healthy stock, 2nd to 5th brood, showing no signs of stress such as mortality >20%, presence of males, ephippia or discoloured animals and there was no delay in the production of the first brood.

CULTIVATION
- Start of each batch: With newborn daphnids, i.e. less than 3 days old, by placing about 250 of them into 5 litres of medium in an all-glass culture vessel.
- Maximum age of the cultures: 4 weeks
- Renewal of the cultures: After 7 days of cultivation half of the medium twice a week.
- Temperature of medium: 18-22°C
- Feeding: Daily, a suspension of fresh water algae.
- Culturing medium: M7

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h

Test conditions

Hardness:
180 mg/L expressed as CaCO3
Test temperature:
20-21°C
pH:
7.6-8.0
Dissolved oxygen:
8.0-9.3 mg/L
Nominal and measured concentrations:
Nominal: 10, 18, 32, 56, 100 mg/L
Measured concentrations: 5.7, 11, 20, 34, 64 mg/L (average). Measured concentrations were 72-78% of nominal at the start of the test. During the exposure period, the concentrations decreased to 44-56% of nominal at the end of the test. Based on these results, the average exposure concentrations were calculated and used to express effect parameters. See Table 1 in 'Any other information on results incl. tables' for details on measured concentrations throughout the test and Table 2 for calculated average exposure concentrations.
Details on test conditions:
TEST SYSTEM
- Test vessel: 60 mL all-glass, containing 50 mL of test solution
- Aeration: no
- Feeding: no
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Adjusted ISO medium, prepared with tap water purified by Reverse Osmosis
- Culture medium different from test medium: yes, M7 medium
- Intervals of water quality measurement: Dissolved oxygen and pH: at beginning and end of the test, for all concentrations and the control. Temperature: continuously in a temperature control vessel, beginning at the start of the test.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h photoperiod daily

EFFECT PARAMETERS MEASURED
- Immobility (including mortality) at 24 hours and at 48 hours.

COMBINED LIMIT/RANGE-FINDING STUDY
- Test concentrations: Control, 0.10, 1.0, 10, 100 mg/L
- Results used to determine the conditions for the definitive study: yes; expected EC50 was between 10 and 100 mg/L (nominal concentration).
Reference substance (positive control):
yes
Remarks:
Potassium dichromate (performed July 2018)

Results and discussion

Effect concentrationsopen allclose all
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
21 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% c.i.: 19-24 mg/L
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
28 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% c.i.: 24-31 mg/L
Details on results:
- Toxicological findings: Immobility increased from 0% at 11 mg/L to 100% at 34 mg/L, at the end of the test.
- Behavioural abnormalities: none observed in the Control group.
- Immobility of control: 5% throughout the exposure period.
- Mortality of control: none
- All water quality parameters remained within the requirements as laid down in the study plan throughout the test.
Results with reference substance (positive control):
- Results with reference substance valid? yes
- The 48h-EC50 was 0.59 mg/L with a 95% confidence interval between 0.53 and 0.72 mg/L.
- The actual responses in this reference test with K2Cr2O7 are within the ranges of the expected responses at the different concentrations, i.e. the 48h-EC50 was between 0.3 and 1.0 mg/L. Hence, the sensitivity of this batch of D. magna was in agreement with the historical data collected at the Test Facility.
Reported statistics and error estimates:
The 24 and 48h-EC50-values were calculated from the weibits of the percentages of affected daphnids and the logarithms of the corresponding test item concentrations (average exposure concentrations) using the maximum likelihood estimation method.
ToxRat Professional v 3.2.1 (ToxRat Solutions® GmbH, Germany) was used to perform the analysis.

Any other information on results incl. tables

Table 1: Average Exposure Concentration Versus Nominal Concentration

 

Test substance, Nominal conc. (mg/L)

Measured concentrations (mg/L)

Average exposure conc. (mg/L)

t=0h

t=48h

10

7.3

4.4               

5.7

18

14

8.3

11

32

25

16

20

56

40

29

34

100

73

56

64

Table 2: Number of Introduced Daphnids and Incidence of Immobility in the Final Test

 

Time (h)

Replicate

Test substance
Average exposure conc. (mg/L)

Control

5.7

11

20

34

64

0

A

5

5

5

5

5

5

B

5

5

5

5

5

5

C

5

5

5

5

5

5

D

5

5

5

5

5

5

Total introduced

20

20

20

20

20

20

24

A

0

0

0

2

     5 (1)

 5*

B

0

0

0

1

5

5

C

0

0

0

     0 (4)

     3 (4)

5

D

1

0

0

     1 (1)

3

5

Total immobilised

1

0

0

4

16

20

Effect %

5

0

0

20

80

100

 

 

 

 

 

 

48

A

0

0

0

2

5

5

B

0

0

0

2

5

5

C

0

0

0

     3 (2)

     5 (2)

5

D

1

0

0

1

     5 (1)

5

Total immobilised

1

0

0

8

20

20

Effect %

5

0

0

40

100

100

( ) between brackets: number of daphnids observed trapped at the surface of the test solutions. These organisms were reimmersed into the respective solutions before recording of mobility.

* Oxygen concentration was 8.9 mg/L.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
For details on validity criteria, please refer to section 'Overall remarks'
Conclusions:
The 48h-EC50 for Daphnia magna exposed to the test item was 21 mg/L, based on average exposure concentrations (95% confidence interval between 19 and 24 mg/L).
Executive summary:

In a 48 h acute toxicity test conducted according to OECD guideline 202, Daphnia magna were exposed to the test item at nominal concentrations of 10, 18, 32, 56 and 100 mg/L and an untreated control (all with 4 replicates per concentration and 5 daphnids per replicate). Measured concentrations decreased to 44 - 56% relative to nominal at the end of the test. Average exposure concentrations were calculated to be 5.7, 11, 20, 34 and 64 mg/L. Immobility increased from 0% at 11 mg/L to 100% at 34 mg/L at the end of the test. The 48h-EC50 for Daphnia magna exposed to the test item was 21 mg/L, based on average exposure concentrations. The study met all validity criteria and is considered to be reliable without restrictions.